Ovarian carcinoma is one of the most aggressive and metastatic forms of female genital cancer, resulting in a disproportionately high death rate compared to incidence. Although epidemiological studies have implicated proliferative events associated with ovulation and repeated wound healing, very little is known about the processes regulating proliferation in either normal human ovarian surface epithelium (OSB) or malignant ovarian epithelial cells. One of the major barriers to studies of normal human OSE cells has been the difficulty in obtaining sufficient cell numbers for molecular and biochemical studies. We have used a recently developed line of SV4O T-antigen transfected human OSE cells with increased proliferative potential to study signaling even associated with gene expression and DNA synthesis in the OSB. These cells are phenotypically normal as evidenced by anchorage dependent growth and lack of tumorigenicity in nude mice. Our preliminary results indicate that activation of protein kinase C (pKC) results in a rapid and persistent inhibition of DNA synthesis in OSE cells, and this inhibition is further increased by thapsigargin treatment and the subsequent release of intra cellular calcium. The observed inhibition of DNA synthesis in normal OSB cells is paralleled by an inhibition of cell proliferation. In contrast, DNA synthesis in the SKOV-3 ovarian tumor cell line is less sensitive to the short term effects of PKC activation, and chronic treatment of SKOV-3 cells with the phorbol ester tumor promoter 12-0-tetradecanoyl-13-phorbol acetate CrpA) actually results in an increase in DNA synthesis, probably due to the depletion of active PKC. These observations have led us to hypothesize that PKC is an important modulator of proliferation in OSB cells, and that changes in PKC-associated signal transduction pathways may be causally related to the loss of growth control in transformed cells. We have also observed that PKC activation in OSE cells results in a rapid and sustained induction of JunB mRNA expression, as well as transient inductions of c-Jun and c-Fos mRNA. We will test the hypothesis that PKC-induced changes in JunB protein expression mediate the effects of PKC activation by observing the effects of JunB depletion and overexpression on DNA synthesis in OSB cells. We will also determine the effects of PKC activation on the phosphorylation of Jun and Fos proteins. We will characterize the interactions between PKC and intracellular calcium, and the expression of PKC isoforms in normal and transformed OSE cells. If the results of these studies support our hypothesis of substantial differences in the PKC-associated signaling pathways between normal and malignant cells, we will test the hypothesis of causality by overexpressing the appropriate PKC isoform in transformed cells and observing the effects on tumorigenicity in nude mice. These studies should provide important information about two major signal transduction systems which may contribute to the regulation of proliferation and differentiation in OSE cells.